Intraspecific sexual isolation in Drosophila.

Intraspecific sexual isolation was examined among wild strains of Drosophila malerkotliana, D. parabipectinata and D. pseudoananassae by multiple-choice method in Elens-Wattiaux mating chamber. In D. pseudoananassae, mating between two strains tested was random and isolation estimate was close to one. In one out of 6 crosses, involving geographic strains of D. malerkotliana, there was significant deviation from randomness and isolation estimate remained low which shows non-random (preferential or positive assortative) mating. In D. parabipectinata, the deviation from randomness was statistically significant due to higher number of homogamic matings in three crosses involving wild strains derived from geographically distant places and isolation estimate remained low in these crosses. The results provide evidence for incipient sexual isolation within D. malerkotliana and D. parabipectinata as a result of genetic divergence.     

Behavioral genetics of Drosophila ananassae

Drosophila ananassae has a unique status among Drosophila species because of certain peculiarities in its genetic behavior. The most unusual feature of this species is its relatively high frequency of spontaneous male recombination. The results of studies on non-sexual behavior, such as phototactic responses, eclosion rhythm, and preferences for oviposition and pupation sites, lead us to suggest that this behavior is under polygenic control, with a substantial amount of additive genetic variation. Sexual isolation has been reported in D. ananassae with the degree of such isolation being stronger in isofemale lines than in natural populations. The significant variations seen in the mating propensity of several isofemale strains, inversion karyotypes and wild type strains, the diminishing effects of certain mutations on the sexual activity of males, and the positive responses to selection for high and low mating propensity point to a genetic control of sexual behavior in D. ananassae. Males contribute more to variation and thus are more subject to intrasexual selection than females. There is a positive correlation between sternopleural bristle number, mating propensity and fertility in D. ananassae. This correlation between morphometric traits and mating success suggests that larger flies are more successful in mating than smaller ones. There is also evidence for adaptive plasticity and a trade-off between longevity and productivity in D. ananassae. Rare, specific courtship song parameters that provide males with a mating advantage have also been reported in different geographic strains of D. ananassae. The remating behavior of males and females, sperm displacement, and the bi-directional selection for female remating speed indicate that post-mating behavior in this species may also be under genetic control. The occurrence of size assortative mating further indicates that there is size-dependent sexual selection in D. ananassae.     

Drosophila bipectinata species complex

The Drosophila bipectinata species complex belongs to the ananassae subgroup of the melanogaster species group (Genus Drosophila, Subgenus Sophophora). The members of the complex are: D. bipectinata, D. parabipectinata, D. malerkotliana, and D. pseudoananassae. Of the four species, D. bipectinata is most widely distributed. Females are indistinguishable, but males are distinguishable by their sex-comb teeth number and pattern and by abdominal colouration. Chromosomal inversions have been detected in these species. In natural populations of D. bipectinata the frequency of inversions and the level of inversion heterozygosity were found to be very low but in laboratory stocks inversions persisted for more than 20 generations due to heterotic buffering. On an average 9.3 fixed interspecific inversions separate each species pair. Non-random association between linked inversions indicated epistatic interaction in natural populations of D. bipectinata. Certain spontaneous mutations were detected and mapped for the first time in D. bipectinata. Low frequency of spontaneous male recombination has also been reported in D. bipectinata. Sexual isolation study in the complex indicated strong preference for homogamic mating. The results also indicated incomplete sexual isolation among different members of this complex. The isolation estimate among six different geographic populations of D. bipectinata ranged from 0.54 - 0.92 representing positive assortative mating which is an evidence for incipient sexual isolation. Incipient sexual isolation was also found within D. malerkotliana and D. parabipectinata . Chromosomal, hybridization and allozyme studies revealed close phylogenetic relationship among the four species of the bipectinata complex. Mitochondrial DNA study revealed net nucleotide difference (delta) between these species to be very small (0.0002 +/- 0.0008) reflecting closeness. Evidence for genetic control of sexual activity and existence of sexual selection in D. bipectinata has been shown on the basis of mating propensity tests carried out on geographic strains, their hybrids and diallel crosses. Significant variation was found among the strains tested with respect to courtship time, duration of copulation and fertility. A positive correlation between duration of copulation and fertility in D. bipectinata was found. Evidence for rare-male mating advantage was also found in D. bipectinata. A positive response to selection for high and low mating activity provided evidence for polygenic control of this phenomenon in D. bipectinata. Bilateral outgrowths on thorax, a unique phenotype, reported for the first time in D. bipectinata has been shown to affect mate recognition ability. Results of the study on pupation site preference (larval behaviour) and oviposition site preference (non-sexual behaviour) have also been included.     

Search for species-specific mating signal in courtship songs of sympatric sibling species,Drosophila ananassae and D. pallidosa

Sexual isolation is one of the most important mechanisms that may lead to speciation. Drosophila ananassae and D. pallidosa are useful for the study of sexual isolation, because of their sympatric distribution and no postmating isolation between them. Courtship songs are considered to play a crucial role in sexual isolation between D. ananassae and D. pallidosa. We recorded and analyzed male courtship songs of D. ananassae and D. pallidosa for eight and four geographical strains, respectively. Courtship behaviors of the two species were consistent with those previously described, however, male's middle leg shaking, which had not before been described, was observed in both species. Males sing by wing vibration only during courtship. Their song oscillograms were distinct between species, but those of conspecific strains were very similar, in spite of their different geographical derivation. We found species-specificity in burst length, pulse length, cycle number in a pulse, and frequency spectra of bursts; these results suggest that these song parameters may play a role in mate recognition that enforces their sexual isolation. The specific values of interpulse interval, cycle number in a pulse and intrapulse frequency were involved with the determination of specificity in frequency spectra of bursts. We discussed the possibility that the specific frequency spectra of bursts are recognized by females as the species-specific signal rather than each parameter individually.     

Effect of temperature on behavioural isolation: a study with Drosophila ananassae populations

Experiments were conducted to study sexual isolation among two natural populations of Drosophila ananassae maintained at 18 degrees C and 24 degrees C for 12 generations in the laboratory to see the effect of this environmental variable on behavioural isolation. Multiple choice technique was used and matings were observed directly in Elens Wattiaux mating chamber. Results showed sexual isolation among strains that were maintained at different temperatures, indicating that temperature may have affected the mating behaviour of the flies which resulted in the induction of ethological isolation among the strains.     

Interspecific sexual isolation and phylogeny among different members of the Drosophila bipectinata species complex

The degree and pattern of sexual isolation among closely related species can be utilized for predicting the direction of evolution and deriving a phylogeny. The Drosophila bipectinata species complex is a group of four morphologically very similar species, belonging to the ananassae subgroup of the melanogaster species group. Sexual isolation among the members of this complex was studied by employing the male-choice technique. To test the difference between the homogamic and heterogamic matings, chi-square values were calculated under the assumption of random mating and the degree of sexual isolation was measured by estimating isolation index. To test the difference in proportions of heterogamic matings between reciprocal crosses, normal deviate (z) was calculated. The direction of evolution among these species has been discussed in the light of models proposed by different workers. Our results suggest that while D. bipectinata, Drosophila parabipectinata and Drosophila malerkotliana are closely related to each other, D. pseudoananassae is distantly related to these three species. The species pairs D. bipectinata and D. parabipectinata and D. parabipectinata and D. malerkotliana show asymmetrical sexual isolation. Based on this, it is suggested that D. bipectinata and D. malerkotliana share a common ancestor where as D. parabipectinata has been derived from D. bipectinata. Their phylogenetic relationship has been discussed in the light of phylogenies suggested by earlier workers.     

No character displacement for reproductive isolation between Drosophila bipectinata and Drosophila malerkotliana.

To test whether character displacement for reproductive isolation between Drosophila bipectinata and Drosophila malerkotliana exists, the degree of sexual isolation was measured between their sympatric and allopatric populations. Although the isolation indices vary in different crosses, the average isolation index for sympatric populations is very close to that for allopatric populations. This shows no difference in the degree of sexual isolation between sympatric and allopatric populations of D. bipectinata and D. malerkotliana. Thus there is no evidence for the existence of character displacement for sexual isolation between these two closely related sympatric species.     

Origin of sexual isolation in <Emphasis Type="Italic">Drosophila ananassae</Emphasis> due to founder effects

The origin of sexual isolation is the central event in the evolution of biological species and plays a key role in maintaining biological diversity. Three mass culture stocks of D. ananassae originating from different geographic localities showing no isolation with each other were subjected to different degrees of bottlenecks i.e. one pair, five pairs and ten pairs. These drift lines were passed through flush-crash cycle at every generation with same initial number of founders, and maintained for twenty-seven generations and then the pattern of matings was tested among these nine drift lines involving 36 crosses in total. In 23 of 36 crosses, the difference between homogamic and heterogamic matings was significant and isolation indices were significantly more than zero in one direction only providing evidence for asymmetrical sexual isolation. Further, when Bonferroni test for pair-wise analysis was employed, significant differences between homogamic and heterogamic matings were found in 25 crosses. These findings provide evidence for origin of sexual isolation by founder effects in D. ananassae.     

Mating activity and fitness of a few wild type strains of Drosophila ananassae

Courtship time, duration of copulation and fertility were tested in six wild type strains of D. ananassae originating from different geographical localities. The results indicate that there is significant variation among the strains tested with respect to courtship time, duration of copulation and fertility. The strains showing a longer duration of copulation produce more progeny. These findings suggest that there is a positive correlation between duration of copulation and fertility in D. ananassae.     

Effect of chromosome arrangements on mate recognition system leading to behavioral isolation in <Emphasis Type="Italic">Drosophila ananassae</Emphasis>

The mechanisms of speciation that appear in the early stages of reproductive isolation has been of recent interest to evolutionary biologists. Experiments were conducted to study behavioral isolation between karyotypically different homozygous strains derived from natural populations of Drosophila ananassae. Three mass cultures stocks established from flies collected from natural populations were employed and homozygous stocks (ST/ST and AL/AL) were made through selection for homozygosity. By employing male-choice technique, mating success was scored by direct observation in the Elens–Wattiaux mating chamber. There is preference for homogamic matings in all the three populations and the differences between homogamic and heterogamic matings are statistically significant in two populations (Lucknow and Varanasi). These findings provide evidence that there is incipient sexual isolation between karyotypically different strains of D. ananassae derived from natural populations which shows that chromosome arrangements may affect the mate recognition system in D. ananassae.     

THE GENETIC BASIS OF SEXUAL ISOLATION BETWEEN DROSOPHILA MELANOGASTER AND D. SIMULANS

The genetic analysis of sexual isolation between the closely-related species Drosophila melanogaster and Drosophila simulans involved two experiments with no-choice tests. The efficiency of sexual isolation was measured by the frequency of courtship initiation and interspecific mating. We first surveyed the variation in sexual isolation between D. melanogaster strains and D. simulans strains of different geographic origin. Then, to investigate variation in sexual isolation within strains, we made F₁ diallel sets of reciprocal crosses within strains of D. melanogaster and D. simulans. The F₁ diallel progeny of one sex were paired with the opposite sex of the other species. The first experiment showed significant differences in the frequency of interspecific mating between geographic strains. There were more matings between D. simulans females and D. melanogaster males than between D. melanogaster females and D. simulans males. The second experiment uncovered that the male genotypes in the D. melanogaster diallel significantly differed in interspecific mating frequency, but not in courtship initiation frequency. The female genotypes in the D. simulans diallel were not significantly different in courtship initiation and interspecific mating frequency. Genetic analysis reveals that in D. melanogaster males sexual isolation was not affected by either maternal cytoplasmic effects, sex-linked effects, or epistatic interaction. The main genetic components were directional dominance and overdominance. The F₁ males achieved more matings with D. simulans females than the inbred males. The genetic architecture of sexual isolation in D. melanogaster males argues for a history of weak or no selection for lower interspecific mating propensity. The behavioral causes of variation in sexual isolation between the two species are discussed.     

THE POTENTIAL FOR GENETIC EXCHANGE BY TRANSFORMATION WITHIN A NATURAL POPULATION OF BACILLUS SUBTILIS

We have investigated the potential for genetic exchange by transformation within a Mojave Desert population of Bacillus subtilis. Almost all strains surveyed were competent for transformation, and the strains varied over almost three orders of magnitude in their levels of competence. This high degree of variation suggests that natural selection toward an optimal level of competence is, at most, very weak in this population. Six of 24 competent strains showed sexual isolation from laboratory strain 168 (i.e., heterogamic transformation was reduced). Direct crosses between selected pairs of Mojave strains indicated sexual isolation within the Mojave population. Levels of sexual isolation observed within this population of B. subtilis were much less than those previously observed for transformation between named Bacillus species. Sexual isolation between 168 and Mojave strains, and among Mojave strains, was due to differences in restriction-modification systems and to DNA sequence divergence.     

Intra- and interspecies variations in pupation height in Drosophila.

In order to study the pupation site preference which is an important component of larval behaviour, pupation height was scored in Drosophila ananassae, D. bipectinata and D. malerkotliana by using a large number of strains of these species. The mean pupation height of different strains ranged from 1.1 to 8.7 mm in D. ananassae, from 0.41 to 0.75 mm in D. bipectinata and from 1.0 to 1.3 mm in D. malerkotliana. The analysis of variance and t-test were performed to test intre- and interspecies variations in pupation height. These tests revealed significant variation among three species. Significant variations among different strains of the same species were also found in D. ananassae and D. bipectinata. These observations provide evidence for intra- and interspecies variations in pupation height in Drosophila. Variations among different strains of the same species in pupation height are attributable to genetic heterogeneity among the strains.     

Wolbachia Infection and Cytoplasmic Incompatibility in Drosophila Species

Forty-one stocks from 30 Drosophila species were surveyed for Wolbachia infection using PCR technology. D. sechellia and two strains of D. auraria were found to be infected and were tested for the expression of cytoplasmic incompatibility, along with D. ananassae and D. melanogaster strains, which are already known to be infected. D. ananassae and D. melanogaster show levels of incompatibility up to 25%, while D. auraria and D. sechellia exhibit levels of egg mortality ~60%. A dot-blot assay using the dnaA sequence as probe was developed to assess the infection levels in individual males that were used in incompatibility crosses. A positive correlation between bacterial density and cytoplasmic incompatibility was observed. The stocks examined can be clustered into at least two groups, depending on the levels of infection relative to the degree of cytoplasmic incompatibility exhibited. One group, containing D. simulans Hawaii, D. sechellia, and D. auraria, exhibits high levels of cytoplasmic incompatibility relative to levels of infection; all the other species and D. simulans Riverside exhibit significantly lower levels of cytoplasmic incompatibility relative to levels of infection. These data show that, in addition to bacterial density, bacterial and/or host factors also affect the expression of cytoplasmic incompatibility.     

Potential gene flow in natural populations of the Drosophila ananassae species cluster inferred from a nuclear mitochondrial pseudogene

A pseudogene with 94% similarity to mitochondrial cytochrome c oxidase subunit I (COI) was identified and localized to chromosome 4 of Drosophila ananassae. Because this chromosome is believed to have reduced recombination, its history can be traced using the pseudo-COI sequence. Pseudo-COI sequences were obtained from 27 iso-female lines of six taxa belonging to the D. ananassae species cluster in which reproductive isolation is incomplete. The phylogenetic network constructed from seven recognized haplotypes (#0-#6) indicated that different taxa inhabiting the same geographic area share the haplotypes: #1 from Papua New Guinean populations of D. ananassae and pallidosa-like-Wau; #2 from Papua New Guinean populations of D. ananassae, pallidosa-like, and papuensis-like; and #4 from South Pacific populations of D. ananassae and D. pallidosa. Taxon-K has a unique haplotype (#6), and 18 mutation steps separate it from the closest haplotype, #2. We discuss the possibility of chromosome 4 introgression beyond taxon boundaries.     

Disruptive selection for sternopleural bristle phenotypes in Drosophila ananassae

Disruptive selection is potentially critical in maintaining variation and initiating speciation and plays an important role in the organization of genetic variability in natural populations. It occurs when extreme phenotypes have a fitness advantage over intermediate phenotypes. Disruptive selection for high and low numbers of sternopleural bristles in Drosophila ananassae was applied for 12 generations to test its effect in induction of behavioural isolation. Pattern of mating between flies of high and low lines was tested in an Elens-Wattiaux mating chamber by using a multiple-choice technique after G(5) and G(12). Data was analyzed by calculating χ(2) under the assumption of random mating to test the difference between homoand heterogamic matings, and sexual isolation was tested by calculating the isolation estimate. The results show that there is no evidence for sexual isolation in G(5) and G(12). The realized heritability, standard error of regression coefficient, and t values suggest that disruptive selection for sternopleural bristle phenotypes was effective, but it does not lead to behavioral isolation in D. ananassae. Rather, it induces differences in mating propensity, which is influenced by sternopleural bristle phenotypes.     

Male Remating in Drosophila ananassae: Evidence for Interstrain Variation in Remating Time and Shorter Duration of Copulation during Second Mating

In Drosophila ananassae, male remating was studied using ten mass culture stocks which were initiated from flies collected from different geographic localities. Male remating occurs at a high frequency and varies within narrow limits (84-96 percent) in different strains. Interestingly, male remating time (in min) varies from 7.41 (Bhutan) to 21.59 (PAT) in different strains and the variation is highly significant. Further, the results also show that males copulate for shorter duration during second mating. This is the first report in the genus Drosophila which provides evidence for interstrain variations for male remating time as well as for shorter duration of copulation during second mating as compared to first mating in D. ananassae.     

No evidence for minority male mating advantage in wild type strains of Drosophila ananassae tested in multiple-choice experiments

Minority male mating advantage was tested in wild type strains of Drosophila ananassae through multiple-choice experiments. Mating success of two types of flies present in five different ratios was scored by direct observation in an Elens-Wattiaux mating chamber. We found no evidence for minority male mating advantage in wild type strains of D. ananassae. The relative mating success of two types of females was also compared in the multiple-choice experiments at different ratios; there was no evidence for a rare female effect. Further, there was similarity in the results of experiments employing different methods. The total number of homogamic and heterogamic matings was obtained by combining the data (all five ratios) from each experiment. Homogamic matings were significantly more frequent than heterogamic ones, which demonstrates preferential mating between males and females of the same strain; this was also supported by a lower isolation estimate. There was also a significant difference in the degree of mating preference between the two strains.     

Genetic analysis of female mating recognition between Drosophila ananassae and Drosophila pallidosa: application of interspecific mosaic genome lines

Drosophila ananassae and Drosophila pallidosa are closely related species that can produce viable and fertile hybrids of both sexes, although strong sexual isolation exists between the two species. Females are thought to discriminate conspecific from heterospecific males based on their courtship songs. The genetic basis of female discrimination behavior was analyzed using isogenic females from interspecific mosaic genome lines that carry homozygous recombinant chromosomes. Multiple regression analysis indicated a highly significant effect of the left arm of chromosome 2 (2L) on the willingness of females to mate with D. ananassae males. Not only 2L but also the left arm of chromosome X (XL) and the right arm of chromosome 3 (3R) had significant effects on the females' willingness to mate with D. pallidosa males. All regions with strong effects on mate choice have chromosome arrangements characterized by species-specific inversions. Heterospecific combinations of 2L and 3R have previously been suggested to cause postzygotic reproductive isolation. Thus, genes involved in premating as well as postmating isolation are located in or near chromosomal inversions. This conclusion is consistent with the recently proposed hypothesis that "speciation genes" accumulate at a higher rate in non-recombining genome regions when species divergence occurs in the presence of gene flow.     

Genetic analyses of several Drosophila ananassae-complex species show a low-frequency major gene for parthenogenesis that maps to chromosome 2

Parthenogenetic strains of several species have been found in the genus Drosophila. The mode of diploidization in the eggs of females has been found to be post-meiotic nuclear fusion. The genetic basis for this parthenogenesis is not understood but is believed to be under the control of a complex polygenic system. We found parthenogenetic females in an isofemale strain (LAE345) of D. pallidosa-like collected in 1981 at Lae, Papua New Guinea, and established a parthenogenetically reproducing strain. Parthenogenetic strains of D. ananassae and D. pallidosa collected at Taputimu, American Samoa had also been established by Futch (1972). D. ananassae, D. pallidosa and D. pallidosa-like are very closely related species belonging to the ananassae complex of the ananassae species subgroup of the melanogaster species group. Using these three species, we found that more than 80% of females from parthenogenetic strains produced progeny parthenogenetically and that inter-specific hybrid females also produced impaternate progeny. In the present report, we demonstrate that the mode of parthenogenesis of D. ananassae appears to be the post-meiotic nuclear doubling of a single meiotic product, and that a major gene responsible for the parthenogenesis maps to the left arm of the second chromosome of D. ananassae. We also suggest that the genetic basis for parthenogenesis capacity may be identical among the three closely related species. We discuss the function of the gene required for parthenogenesis and its significance for the evolutionary process.